Allergen description

Fel d 4, a protein of approximately 20 kDa in size, belongs to the lipocalin protein family.

In a study of the frequency of IgE reactivity to recombinant Fel d 4 in 27 allergic subjects tested, 62.96% (17 of 27) had detectable serum IgE to this allergen. This was similar to the frequency of IgE reactivity detected for rFel d 1 in the same population (88.88%; 24 of 27). Of 16 non-allergic controls, two subjects were borderline positive to rFel d 4; both of these subjects were skin-prick test-positive to cat but had no clinical symptoms. (1) However, although a high frequency of cat-allergic subjects were found to have measurable IgE antibody to Fel d 4, typically the levels were low, i.e. Fel d 4 appears to have frequent but quantitatively low binding of IgE from the sera of cat-allergic people. (1)

In 1987 researchers reported that a 22 kDa protein in cat extract had a high frequency of IgE-binding activity. (2) It is feasible that this may correspond to natural Fel d 4. (1) Other studies have suggested that there is a signi?cant group of cat-allergic individuals who have minimal anti-Fel d 1 IgE reactivity. (25, 26) The assumption has been that cat allergy is almost entirely caused by reactivity to the major Fel d 1 protein; however, recent data suggest that other allergens such as Fel d 4 may play some role in sensitisation to the domestic cat. (1)

Lipocalins have been detected in urine, tears, saliva, serum and dander across a wide range of species. However, Fel d 4 has been isolated from the submandibular salivary gland of the cat, and analysis has shown that the mRNA for the allergen was limited to this tissue and was not found in the parotid gland, the liver, skin, the tongue or the anal gland.1

The degree of sequence identity between lipocalins is usually low, often in the order of 10-20%; however, Fel d 4 has a high amino acid sequence identity with (and is highly homologous with) several other mammalian lipocalins: to the major horse allergen Equ c 1 (67.3%), and a lower level of identity to boar salivary lipocalin (Sal 1, 62%), murine MUP (Mus m 1, 48%) and rat a-2m globulin (Rat n 1, 53.6%). (1) The level of identity to other lipocalin allergens is much lower, e.g. Can f 1 (21%) and Bos d 2 (26%). (1) The allergenicity of boar Sal 1 is unknown.

The lipocalins are a large, diverse group of at least 50 proteins. Lipocalins are a large protein group comprising small extracellular proteins present in vertebrate and invertebrate animals, plants and bacteria. Lipocalins are proteins that are present in humans, and it is notable that almost all important respiratory allergens from mammals belong to this protein family. (27)

Lipocalins are typically small, acidic glycoproteins present in body fluids and secretions, produced by the liver or secretory glands. They share common biological functions, predominantly related to the transport of small hydrophobic molecules, such as odorants, steroids, vitamins and pheromones; i.e. their most prominent feature is their ability to bind small hydrophobic molecules such as steroids. Although they were originally characterised as transport proteins for diverse molecules (such as odorants, steroids, and pheromones), they are involved in a wide range of other biological functions. (28) Some lipocalins show immunomodulatory activity. ß-lactoglobulin (Bos d 5) and tear lipocalin have been reported to have nonspecific endonuclease activity. (28)

Several lipocalins are able to bind and transport small hydrophobic ligands such as retinol. (29, 30, 31) Major respiratory allergens of dogs, mice, rats, horses and cows belong to the lipocalin group of proteins. (32) In addition to mammalian respiratory allergens, the milk allergen Bos d 5 (ß-lactoglobulin), the cockroach allergen (Bla g 4), a "kissing bug" (Triatoma protracta) allergen (Tria p 1), (33) and a pigeon tick (Argas reflexus) allergen (Arg r 1) (34) all belong to the lipocalin group. (28)

Several lipocalins have been described as allergenic proteins, including: (35)

Mouse

Mus m 1

18-21 kDa

mouse major urinary protein mMUP

Rat

Rat n 1

17-21 kDa

rat urinary a2-globulin

Cow

Bos d 2

Bos d 5

bovine dander lipocalin

bovine milk ß-lactoglobulin

Cockroach

Bla g 4

cockroach lipocalin

Dog

Can f 1

Can f 2

dog lipocalin

dog lipocalin

Horse

Equ c 1

Equ c 2

horse lipocalin

horse lipocalin

Guinea pig

Cav p 1

Cav p 2

guinea pig lipocalin

guinea pig lipocalin

Cat

Fel d 4

cat lipocalin

Rabbit

Ory c 1

Ory c 2

rabbit lipocalin

rabbit lipocalin

Lipocalins are members of the calycin superfamily. Despite the diversity at the primary sequence level and a sequence identity often less than 20%, all lipocalins share a conserved folding pattern, a similar three-dimensional structure and one to three conserved regions: an 8-stranded ß-barrel flanked by an a-helix at the C-terminal end of the polypeptide chain. (29) The central cavity of the lipocalin ß-barrel serves for the binding and transport of small hydrophobic molecules such as retinol (retinol-binding protein), odorant molecules (bovine odorant-binding protein), or – as in mouse and rat – for pheromones (mouse major urinary protein, mMUP1, and rat urinary a2-globulin). (35)

Although the overall amino acid identity between lipocalins is usually below 20%, in some cases the sequential identity over animal species can be well above 20%. For example, Fel d 4 has a high amino acid sequence identity with the major horse allergen Equ c 1 (67.25%),1 and dog Can f 1 exhibits a 57% identity with human tear lipocalin (Von Ebner's gland protein), and human epididymal-specific lipocalin-9 exhibits a 40% to 50% identity with rodent lipocalins. (28) Lipocalins exist as both monomers and dimers, and they can be either glycosylated or nonglycosylated. (28)

Immune reactivity to lipocalin allergens is not completely understood. Although the cellular immune response to lipocalin allergens has not yet been fully characterised, in general it appears to be weak. Researchers have noted that this is surprising, as humans mount a strong IgE response against these allergens; and therefore it was proposed that the presence of endogenous lipocalins might be a factor contributing to the T-helper type 2 (Th2) deviation of the immune response against exogenous lipocalin allergens. (27)

Other researchers have argued that in Bos d 5 the IgE-binding epitopes are spread along the molecule, whereas in Bos d 2 the C-terminus appears to contain the human B cell epitopes; and that Bos d 5 contains several murine T-cell epitopes. They therefore propose that the allergenicity of lipocalins may be a consequence of molecular mimicry between lipocalin allergens and endogenous lipocalins at the T-cell level. (36)

Other researchers have argued that the allergenicity of lipocalin is a consequence of molecular mimicry between endogeneous lipocalins and exogenous lipocalin allergens at the T-cell level. (37)

Despite the low amino acid sequence identity often being less than 20%, and the diversity at the primary sequence level, as all lipocalins share a conserved folding pattern cross-reactivity between lipocalins is possible. (29)

However the clinical significance of IgE cross-reactivity between mammalian non-serum-derived respiratory allergens remains unclear. (29) The cross-reactivity appears to be mostly weak to moderate. Earlier analyses were based on inhibiting IgE binding to an allergen extract by another extract: the inhibition was usually found to be individually variable. In addition, the extracts often showed an unequal inhibitory capacity, suggesting that only parts of the IgE-binding epitopes were common. (29)

A number of studies have shed some light on possible cross-reactivity between Fel d 4 and other lipocalin allergens.

For example, it has been demonstrated that the greatest degree of cross-reactivity was between dander extracts of cats and dogs, and not saliva, which – given the origin of Fel d 4 cDNA in the salivary gland – could be a major source of the secreted allergen. (38) Yet there appears to be no (or very little) cross-reactivity between Fel d 4 and dog dander extract. (1)

A study describing the characterisation of Fel d 4, a cat dander lipocalin, reported high sequence identity to the boar salivary lipocalin and the horse lipocalin Equ c 1. IgE binding to Fel d 4 could be blocked by an allergen extract from cow, and to a lesser degree by extracts from horse and dog. (1)

Asthma, occupational asthma, allergic rhinitis and allergic conjunctivitis are commonly induced by cat allergens. (39, 40) Domestic animals represent the second-most important group of indoor air allergens after house dust mites. (41) Sensitisation may occur even following unobserved exposure to cat, or where the animal is no longer present, due to the persistence of the allergen; the allergen may be brought in on visitors' clothing. (42, 43) Cat dander allergens have even been found in dust storms. (44)

See Cat dander e1 for clinical information and further details on Cat dander allergy. See also Cat serum albumin nFel d 2 e220.

Some studies suggest that those allergic to cat dander have a significantly lower risk of current asthma than those not allergic to cat dander and not owning a cat, but these findings have been contradicted by other reports. (45) Sensitivity to dog or cat dander or Alternaria, as determined by skin-specific IgE, was shown to be associated with increased bronchial responsiveness but not with decreased lung function in children with mild to moderate asthma. These findings support the important role that sensitisation to certain allergens plays in modulating bronchial responsiveness. (46)

Studies throughout the world, wherever cats are common domestic pets, report high sensitisation to cat. (47, 48, 49, 50)

Recombinant allergens, which are genetically engineered isoforms resembling allergen molecules from known allergen extracts, have immunoglobulin E (IgE) antibody binding comparable to that of natural allergens, and generally show excellent reactivity in in vitro and in vivo diagnostic tests. (51 )To date, many different recombinant allergens of various pollens, moulds, mites, and foods – as well as latex and bee venom – have been cloned, sequenced, and expressed.

Recombinant allergens have a wide variety of uses, from the diagnosis and management of allergic patients, to the development of immunotherapy, to the standardisation of allergenic test products as tools in molecular allergology. Recombinant allergens are particularly useful for investigations of allergies manifesting wide cross-reactivity.

As the number of important allergens in commercial cat extracts can vary extensively, and as natural preparations may be contaminated with other components, potentially causing false-positive skin-specific IgE test results, recombinant Fel d 4 has a role to play in assessing allergy to cat.

Importantly, cat dander-allergic individuals are sensitised to a heterogenous range of cat allergens. Recombinant allergens enable assessment of sensitisation to specific allergens in the repertoire; and specifically, Fel d 4 provides a means for investigating differences in the immune response to lipocalin allergens from those found for other mammalian species, but is also an important reagent for the diagnosis of cat allergy. (1)

Panzani RC, Mercier P, Delord Y, Riva G, Falagiani P, Reviron D, Auquier P. Prevalence of patent and latent atopy among a general normal adult population in the south east of France by RAST investigation and correlation with circulating total IgE levels. Allergol Immunopathol (Madr) 1993;21(6):211-9.

Perzanowski MS, Ronmark E, Nold B, Lundback B, Platts-Mills TA. Relevance of allergens from cats and dogs to asthma in the northernmost province of Sweden: schools as a major site of exposure. J Allergy Clin Immunol 1999;103(6):1018-24.